TY - JOUR
T1 - Development of a method for quantification of acrolein-deoxyguanosine adducts in DNA using isotope dilution-capillary LC/MS/MS and its application to human brain tissue
AU - Liu, Xinli
AU - Lovell, Mark A.
AU - Lynn, Bert C.
PY - 2005/9/15
Y1 - 2005/9/15
N2 - Acrolein is a highly reactive α,β-unsaturated aldehyde and is known to react with DNA forming exocyclic acrolein-deoxyguanosine adducts (Acro-dG). These aldehyde-DNA lesions may play a role in mutagenesis, carcinogenesis, and neurodegenerative diseases. In the present work, we described the development and evaluation of a highly sensitive and selective capillary liquid chromatography nanoelectrospray isotope dilution tandem mass spectrometry method for quantitatively analyzing Acro-dG in DNA hydrolysates. This was achieved by applying a stable isotope-labeled analogue Acro-dG- 13C10, 15N5 as an internal standard to the DNA to be analyzed and then hydrolyzing the DNA enzymatically to nucleosides. The acrolein-modified nucleosides were separated from normal nucleosides by capillary liquid chromatography and quantified by a high-capacity ion trap mass spectrometer in the MS/MS mode. The developed method achieved attomole-level sensitivity (limit of detection was 10 fg, 31 amol on column) for detection of pure Acro-dG adduct standards. The limit of quantification of Acro-dG adducts obtained in 10 μg of DNA hydrolysates was 1.5 fmol, which corresponded to 50 adducts/109 normal nucleosides. Application of this method to the analysis of Acro-dG adducts in acrolein (10-fold)-treated calf thymus DNA showed ∼830 lesion/106 DNA nucleosides using as low as 50 ng of DNA. Application of this method to DNA samples (1-2 μg) isolated from brain tissues from Alzheimer's disease subjects and age-matched controls demonstrated 2800-5100 Acro-dG adducts/109 DNA nucleosides. Statistically significant differences (P < 0.05) in levels of Acro-dG between AD subjects and controls were observed in DNA isolated from the hippocampus/ parahippocampal gyrus.
AB - Acrolein is a highly reactive α,β-unsaturated aldehyde and is known to react with DNA forming exocyclic acrolein-deoxyguanosine adducts (Acro-dG). These aldehyde-DNA lesions may play a role in mutagenesis, carcinogenesis, and neurodegenerative diseases. In the present work, we described the development and evaluation of a highly sensitive and selective capillary liquid chromatography nanoelectrospray isotope dilution tandem mass spectrometry method for quantitatively analyzing Acro-dG in DNA hydrolysates. This was achieved by applying a stable isotope-labeled analogue Acro-dG- 13C10, 15N5 as an internal standard to the DNA to be analyzed and then hydrolyzing the DNA enzymatically to nucleosides. The acrolein-modified nucleosides were separated from normal nucleosides by capillary liquid chromatography and quantified by a high-capacity ion trap mass spectrometer in the MS/MS mode. The developed method achieved attomole-level sensitivity (limit of detection was 10 fg, 31 amol on column) for detection of pure Acro-dG adduct standards. The limit of quantification of Acro-dG adducts obtained in 10 μg of DNA hydrolysates was 1.5 fmol, which corresponded to 50 adducts/109 normal nucleosides. Application of this method to the analysis of Acro-dG adducts in acrolein (10-fold)-treated calf thymus DNA showed ∼830 lesion/106 DNA nucleosides using as low as 50 ng of DNA. Application of this method to DNA samples (1-2 μg) isolated from brain tissues from Alzheimer's disease subjects and age-matched controls demonstrated 2800-5100 Acro-dG adducts/109 DNA nucleosides. Statistically significant differences (P < 0.05) in levels of Acro-dG between AD subjects and controls were observed in DNA isolated from the hippocampus/ parahippocampal gyrus.
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U2 - 10.1021/ac050624t
DO - 10.1021/ac050624t
M3 - Article
C2 - 16159131
AN - SCOPUS:24944565214
SN - 0003-2700
VL - 77
SP - 5982
EP - 5989
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 18
ER -